There is an immediate need for a highly safe and efficacious vaccine that can protect against highly pathogenic avian influenza viruses (H5N1). This strain of influenza virus emerged recently in Southeastern Asia producing severe outbreaks in poultry and wildlife species and has occasionally affected humans with high mortality rates. The virus has quickly spread throughout Asia, Europe and Africa and there is a major public concern that further reassortment might create a virus capable of human- to human transmission that could produce the next flu pandemic. In a related poxvirus vector, we have demonstrated the importance of antigen expression levels/patterns and the subsequent effect in generating protective immune responses. This work resulted in the development of novel recombinant poxvirus vaccines that protect against Y. pestis infection (1). Here we propose to adapt and extend these findings to a related poxvirus vector, Modified vaccinia virus Ankara (MVA). MVA offers distinct advantages as a viral vaccine vector for the construction of recombinant influenza vaccines. In this proposal, we will 1) optimize hemmaglutinin (HA) and nucleoprotein (NP) antigen expression from the H5N1 strain by the MVA vector, 2) evaluate the effect that promoters and antigen expression patterns (e.g. intracellular, secreted, membrane-bound) might have on the immune response, and 3) determine the protection conferred by these recombinant viruses against lethal challenge. The specific aims are as follows. 1) Construct and optimize the expression of HA and NP antigens by MVA recombinant viruses. 2) Evaluate the efficacy of recombinant MVA/FLU vaccines for protection against lethal flu challenge in mice; and 3) Evaluate the safety of the recombinant MVA/FLU vaccines in immunodeficient mice. The vaccine construct that safely provides the most complete protection of mice from intranasal influenza challenge will be chosen for further analysis. In subsequent proposals, the candidate MVA/FLU vaccine will be tested in ferrets and/or non-human primates for the ability to protect against influenza challenge and will be prepared for human clinical trials. [unreadable] [unreadable] [unreadable]